This invention is a further development in the detection of radioactive species by high-pressure liquid chromatography.
In particular, this invention relates to the detection of radioactive species in the effluent from a chromatographic column by passing the effluent through a cell packed with a material that is caused to fluoresce by radioactive emissions in the cell.
More particularly, the present invention provides an apparatus and method for obtaining a chromatogram in real time for the radioactivity occurring in a chromatographic column.
Various techniques have been used in the prior art to detect a radioactive species in the effluent from a chromatographic column by monitoring scintillations produced in a fluorescent composition by radioactive emissions from the species to be detected. According to one technique, the effluent from a chromatographic column was mixed with a "scintillation cocktail", and this mixture was then passed through an open tube (i.e., an unpacked flow cell). The "scintillation cocktail" comprised a fluorescent substance such as 2,5-diphenyl oxazole (known as PPO), which was caused to scintillate by the transfer of molecular excitation energy to it from a solvent such as toluene or dioxane that has been excited when bombarded by radioactive emissions. This "scintillation cocktail" technique is discussed in greater detail in The Current status of Liquid Scintillation Counting, edited by E. O. Branson, published by Grune & Stratton, 1970.
A disadvantage of the "scintillation cocktail" technique is that when the column effluent is aqueous, the ratio of the "scintillation cocktail" volume to the effluent volume must typically be in the range from 5:1 to 10:1 in order to counteract the quenching effect of the aqueous solvent upon the scintillation process, and in order to prevent aqueous precipitation of the scintillation material. Furthermore, to accommodate the addition of such a large quantity of "scintillation cocktail" to the effluent, the flow rate of the mixture through the open flow cell downstream of the column must be increased in order to permit the desired flow rate of the effluent through the column. Such an increase in the flow rate through the open flow cell tends to cause laminar flow-band broadening and reduced resolution.
Another disadvantage of the "scintillation cocktail" technique arises from the fact that a pump must be used to deliver the "scintillation cocktail" to the column effluent, thereby complicating and increasing the cost of the apparatus necessary for the monitoring of scintillations in the downstream flow cell. Furthermore, the "scintillation cocktail" material itself is costly; and the mixing of such a large volume of "scintillation cocktail" per unit volume of effluent is not economically feasible in many applications.
The concept of passing the effluent from a chromatographic column through a flow cell packed with a material that scintillates when exposed to radioactive emissions is also known to the prior art. A review of the pertinent prior art in this regard can be found in the specification of co-pending U.S. patent application Ser. No. 953,380, filed on Oct. 23, 1978, assigned to Varian Associates, Inc. A particular fluorescent composition that is suitable as a flow-cell packing for the monitoring of scintillations caused by radioactive emissions from a species in the effluent passing through a flow cell is microparticulate didansyl-N-2-aminoethyl-3-aminopropyl silica, which is claimed in U.S. patent application Ser. No. 953,380.